Phylogenomics and genetic analysis of solvent-producing Clostridium species

Abstract

The genus Clostridium is a large and diverse group within the Bacillota (formerly Firmicutes), whose members can encode useful complex traits such as solvent production, gas-fermentation, and lignocellulose breakdown. We describe 270 genome sequences of solventogenic clostridia from a comprehensive industrial strain collection assembled by Professor David Jones that includes 194 C. beijerinckii, 57 C. saccharobutylicum, 4 C. saccharoperbutylacetonicum, 5 C. butyricum, 7 C. acetobutylicum, and 3 C. tetanomorphum genomes. We report methods, analyses and characterization for phylogeny, key attributes, core biosynthetic genes, secondary metabolites, plasmids, prophage/CRISPR diversity, cellulosomes and quorum sensing for the 6 species. The expanded genomic data described here will facilitate engineering of solvent-producing clostridia as well as non-model microorganisms with innately desirable traits. Sequences could be applied in conventional platform biocatalysts such as yeast or Escherichia coli for enhanced chemical production. Recently, gene sequences from this collection were used to engineer Clostridium autoethanogenum, a gas-fermenting autotrophic acetogen, for continuous acetone or isopropanol production, as well as butanol, butanoic acid, hexanol and hexanoic acid production.

The Industrial Fermentation Process and Clostridium Species Used to Produce Biobutanol

“…A joint LanzaTech-JGI project has added another 270 genomes from the DJ culture collection. Of these, 72% were C. beijerinckii genomes, consisting of 160 genomes from the NCP industrial strain collection and 34 genomes of strains originating from the international culture collection [2,38]. The phylogenetic analysis from the expanded number of genome sequences confirmed that the C. beijerinckii species consists of four distinct subgroups that possibly can be considered as subspecies.…”

“…The three species are closely related phylogenetically and belong together in a separate phylogenetic clade. The members of the species in this clade encode the sol operon in the gene order ald-ctfA-ctfB-adc and lack a pdc gene [38]. As a general observation, all three species appear to perform more effectively on plant-based substrates such as molasses than they do on maize or laboratory media with glucose and/or sucrose as the sole energy source.…”

“…The original C. acetobutylicum strains used to produce solvents from maize and other starch-based substrates were found to belong to a separate phylogenetic clade. The other three saccharolytic species, C. beijerinckii, Clostridium saccharobutylicum, and Clostridium saccharoperbutylacetonicum, used to produce solvents from molasses and other sugar-based substrates were found be closely related and belong together in a separate distantly-related phylogenetic clade [2,38,39].…”

“…A list of C. acetobutylicum and C. beijerinckii strains from her collection giving their origins is provided in Table 1. A second collection of industrial solvent-producing clostridia, referred to as the David Jones (DJ) culture collection, includes many of the industrial strains available from the international culture collections [2,38]. The collection also includes production strains used for the commercial production of solvent by National Chemical Products (NCP) in South Africa [2,12].…”

“…More than 300 genome sequences are now available for the study of these key industrial clostridial species, sourced from both the international culture collections and the DJ culture collection which is now held by LanzaTech. Conversion tables for the DJ strains that were sequenced have been published recently [2,38]. Although the four species of Clostridium used for industrial solvent production differ phylogenetically, they share many similar characteristics.…”